Project Summary/Abstract Anorexia nervosa (AN) is characterized by severely restrictive eating and emaciation, with high rates of morbidity, chronicity and mortality. Current treatments have high rates of relapse and recurrence, thus developing new and improved therapies is a high priority in mental and public health. While AN is highly heritable, identifying risk genes and associated mechanisms on AN pathophysiology has yet to be successful due to the lack of adequate knowledge on how genes interact with non-genetic risk factors. This study aims to fill this major knowledge gap in order to develop better prevention and treatments. We will examine how soluble epoxide hydrolase (sEH), the gene product of an AN susceptibility gene, EPHX2, affects AN psychopathology and outcome using a meal-challenge study design to elucidate the mechanisms by which EPHX2 affects AN. sEH is a key enzyme that affects lipid signaling functions of bioactive metabolites (termed oxylipins). It modulates cellular inflammation by converting anti-inflammatory epoxides of polyunsaturated fatty acids (PUFAs) to pro-inflammatory oxylipins. In our recent study we identified dysregulated lipidomic profiles and elevated in vivo sEH activity (assessed by oxylipin ratios) in AN patients. The increase of the in vivo sEH marker persisted in weight-recovered AN, further supporting our hypothesis that EPHX2 regulates sEH activity that induces a pro-inflammatory oxylipin shift, therefore elevated sEH may contribute not only to AN susceptibility but also illness course and outcome. Testing this hypothesis is important because if found true, sEH activity may serve as a useful biomarker to predict disease outcome and guide treatment development. PUFAs are precursors of oxylipins, therefore PUFAs may affect regulatory efficiency of sEH on oxlipins. AN patients are known for their strong aversion to meat-based fatty foods (rich in ?-6 PUFA), and we have shown that dietary ?-6:?-3 PUFA ratio is decreased in AN patients, suggesting that dysregulation of the EPHX2 lipidomic pathway in AN may be influenced by their PUFA intake. Thus, we will test the hypothesis that in AN patients, the metabolism of oxylipins is perturbed due to an increase in sEH activity, and the degree of perturbation is PUFA-dependent, resulting in increased food aversion, a higher burden of psychopathology, and poor outcome. In Aim 1 we will test if EPHX2 metabolomic pathways are altered in anorexia nervosa through a ?-6 rich breakfast challenge. In Aim 2 we will emphasize the fundamental biochemistry and molecular mechanism of EPHX2 gene by assaying ex vivo sEH activity to assess its association with AN risk. We will also characterize EPHX2 variants to examine the effects of the EPHX2 non-synonymous SNPs (nsSNPs) on ex vivo sEH activity. In Aim 3, we will utilize pre-meal and post-meal AN psychopathologic traits and 3-month outcome data to assess sEH's role in AN illness course and outcome through a longitudinal design. This study is significant because it will provide insights into AN pathogenesis by the use of an innovative meal challenge protocol that will reveal how genetic and dietary factor work in concert to exert effects on AN risk, phenotypes, and outcome. The results of this study will unravel mechanisms by which EPHX2 affects AN, improves our ability to predict disease risk, monitor disease progression and treatment response, and lead to new targets for future nutraceutical and pharmaceutical developments.